Laurence Bordenave

Laser assisted bioprinting of engineered tissue with high cell density and microscale organization

Over this decade, cell printing strategy has emerged as one of the promising approaches to organize cells in two and three dimensional engineered tissues. High resolution and high speed organization of cells are some of the key requirements for the successful fabrication of cell-containing two or three dimensional constructs. So far, none of the available cell printing technologies has shown an ability to concomitantly print cells at a cell-level resolution and at a kHz range speed. We have studied the effect of the viscosity of the bioink, laser energy, and laser printing speed on the resolution of cell printing. Accordingly, we demonstrate that a laser assisted cell printer can deposit cells with a microscale resolution, at a speed of 5 kHz and with computer assisted geometric control. We have successfully implemented such a cell printing precision to print miniaturized tissue like layouts with de novo high cell density and micro scale organization.

Cell-to-cell communication between osteogenic and endothelial lineages: implications for tissue engineering

There have been extensive research efforts to develop new strategies for bone tissue engineering. These have mainly focused on vascularization during the development and repair of bone. It has been hypothesized that pre-seeding a scaffold with endothelial cells could improve angiogenesis and bone regeneration through a complex dialogue between endothelial cells and bone-forming cells. Here, we focus on the paracrine signals secreted by both cell types and the effects they elicit. We discuss the other modes of cell-to-cell communication that could explain their cell coupling and reciprocal interactions. Endothelial cell-derived tube formation in a scaffold and the dialogue between endothelial cells and mesenchymal stem cells provide promising means of generating vascular bone tissue-engineered constructs.

The effect of RGD density on osteoblast and endothelial cell behavior on RGD-grafted polyethylene terephthalate surfaces

Hybrid materials combining polyethylene terephthalate and different types of cells (endothelial and osteoblastic cells) have been developed thanks to the covalent grafting of different densities of RGD containing peptides onto the polymer surface. Biomimetic modifications were performed by means of a three-step reaction procedure: creation of COOH functions, coupling agent grafting and the immobilization of the RGDC peptides. High resolution mu-imager was used to evaluate RGD densities (varying between 0.6 and 2.4 pmol/mm(2)) and has exhibited the stability of the surface grafted peptides when treated in harsh conditions. The efficiency of this route for biomimetic modification of a PET surface was demonstrated by measuring the adhesion of MC3T3 and HSVEC cells and by focal adhesion observation. Results obtained prove that a minimal RGDC density of 1 pmol/mm(2) is required to improve MC3T3 and HSVEC cells responses. Indeed, cells seeded onto a RGDC-modified PET with a density higher than 1 pmol/mm(2) were able to establish focal adhesion as visualized by fluorescence microscope compared to cells immobilized onto unmodified PET and RGDC-modified PET with densities lower than 1 pmol/mm(2). Moreover, the number of focal contacts was enhanced by the increase of RGDC peptide densities grafted onto the material surface. With this study we proved that the density of peptides immobilized on the surface is a very important parameter influencing osteoblast or endothelial cell adhesion and focal contact formation.

Review of 10 years of the clinical use of brain natriuretic peptide in cardiology

Ten years ago brain natriuretic peptide (BNP), the second compound of a family of polypeptide hormones named natriuretic peptides was identified. This peptide has great pathophysiologic importance as a stress-induced cardiac hormone secreted from ventricles, and it rises in several cardiac diseases. It promotes natriuresis and diuresis, acts as a vasodilator, and antagonizes the vasoconstrictor effects of the renin-angiotensin-aldosterone system. The measurement of this peptide in blood by immunoassay has shown promise over the past decade in clinical diagnosis and prognosis. Because heart failure is a major health problem worldwide, BNP is proposed as a biochemical marker that might provide a useful screening test to select patients for further cardiac investigations. Such a hormone assay is inexpensive and available. The implications of BNP in diagnosis, prognosis, and therapy will be reviewed.

Interaction between human umbilical vein endothelial cells and human osteoprogenitors triggers pleiotropic effect that may support osteoblastic function

Osteogenesis occurs in striking interaction with angiogenesis. There is growing evidence that endothelial cells are involved in the modulation of osteoblast differentiation. We hypothesized that primary human umbilical vein endothelial cells (HUVEC) should be able to modulate primary human osteoprogenitors (HOP) function in an in vitro co-culture model. In a previous study we demonstrated that a 3 day to 3 week co-culture stimulates HOP differentiation markers such as Alkaline Phosphatase (ALP) activity and mineralization. In the present study we addressed the effects induced by the co-culture on HOP within the first 48 hours. As a prerequisite, we validated a method based on immuno-magnetic beads to separate HOP from HUVEC after co-culture. Reverse transcription-real time quantitative PCR studies demonstrated up-regulation of the ALP expression in the co-cultured HOP, confirming previous results. Surprisingly, down-regulation of runx2 and osteocalcin was also shown. Western blot analysis revealed co-culture induced down-regulation of Connexin43 expression in both cell types. Connexin43 function may be altered in co-cultured HOP as well. Stimulation of the cAMP pathway was able to counterbalance the effect of the co-culture on the ALP activity, but was not able to rescue runx2 mRNA level. Co-culture effect on HOP transcriptome was analyzed with GEArray cDNA microarray showing endothelial cells may also modulate HOP extracellular matrix production. In accordance with previous work, we propose endothelial cells may support initial osteoblastic proliferation but do not alter the ability of the osteoblasts to produce extracellular mineralizing matrix.

Development of RGD peptides grafted onto silica surfaces: XPS characterization and human endothelial cell interactions.

The attachment of human umbilical vein endothelial cells (HUVECs) on substrates that had been covalently grafted with the cell adhesion peptides Arg-Gly-Asp (RGD) was investigated. This approach was used to provide substrates that are adhesive to cells even in the absence of serum proteins and to cells that have had no prior treatment of the surface with proteins that promote cell adhesion. We wanted to improve control of cellular interactions with cell-adhesive materials by providing fixedly bound adhesion ligands. Silica was examined as a model surface. The peptides were grafted using three different steps: grafting of aminosilane molecules; reaction with a maleimide molecule; and immobilization of cell-binding peptides containing the RGD sequence. The RGD-grafted surface was characterized by X-ray photoelectron spectroscopy (XPS) and contact-angle measurements.

Positron emission tomography in patients with suspected pacing system infections may play a critical role in difficult cases

BACKGROUND A pacemaker recipient may be hospitalized recurrently with an infection of unknown origin despite detailed investigations. OBJECTIVE The purpose of this study was to investigate whether (18)F-fluorodeoxyglucose positron emission tomography/computerized tomography (FDG-PET/CT) scanning has a role in identifying pacing material infection in these difficult cases. METHODS Ten patients who presented with fever of unknown origin despite detailed investigations including transesophageal echocardiography underwent FDG-PET/CT scanning. Identification of increased FDG uptake along a pacing lead prompted the removal of the entire pacing system, whereas in the absence of increased FDG uptake the pacing material was left in place. Forty control pacemaker recipients underwent FDG-PET/CT scanning as part of investigation of malignancy. RESULTS Among the 40 patients in the control group, FDG-PET/CT scanning was normal in 37 (92.5%) patients. Among the 10 patients who presented with suspected pacing system infections, FDG-PET/CT scanning showed increased FDG uptake along a lead in six patients; as a result of this finding, these patients subsequently underwent complete removal of the implanted material. Cultures of the leads were positive in all six patients, confirming involvement of the leads in the infectious process. In the other four patients, the pacing system was left in place without objective signs of active lead endocarditis during follow-up. CONCLUSION This study demonstrates the potential value of FDG-PET/CT scanning in the diagnosis of pacing lead endocarditis in difficult cases. Increased FDG uptake along a lead in this clinical context appears to be a reliable sign of active infection.

Developments towards tissue- engineered, small-diameter arterial substitutes

This article reviews the present state of small-diameter vascular substitution, the mechanisms of graft failure and the ongoing matters of graft design, with particular attention to small-caliber grafts. Tissue engineering of blood vessels with the mechanical properties of native vessels and with the antithrombotic properties required is really a great challenge. Recent advances, however, indicate that the goal of providing a tissue-engineered vascular graft that will remain patent in vivo for substantial periods of time, is achievable. This review describes the specific requirements of the successful replacement artery from a biomaterial- and tissue-engineering perspective, as well as various approaches in vascular tissue engineering including cells and cell sources, scaffolds and their improvements as well as bioreactors and signaling.

New artificial connective matrix-like structure made of elastin solubilized peptides and collagens: elaboration, biochemical and structural properties.

Several improvements of the basic reaction between elastin peptides and type III collagen, specially the addition of heparan sulphate proteoglycans, are presented. The consequent elaboration of either cell culture support or membranes are described and illustrated by scanning electron microscopy. The material produced possesses composition, very well organized lamellar structure and some properties quite close to natural membranes such as subendothelial tissue.

Physico-chemistry and cytotoxicity of ceramics: Part II Cytotoxicity of ceramics

General cytotoxicity was assayed for ceramic (Al2O3, ZrO2/Y2O3, AIN, B4C, BN, SiC, Si3N4, TiB4, TiC, TiN) diamond and graphite powders, using 3T3 Balb/c permanent cell lines. Neutral red test was carried out in order to establish cell viability. Further investigations were undertaken on human differentiated cells (human umbilical venous endothelial cells): cell behaviour (MTT assay, total cell protein content) and differentiation (immunofluorescence) were studied. In both cases, no cytotoxic effect has been noticed. All the impurities contained at low concentration in these powders do not seem to present any effect. The correlation which has been previously observed between cytotoxicity-cell culture response and blood haemolysis for polymers has not been established here for ceramic powders. We conclude that all the ceramic powders tested here and therefore the corresponding bulk ceramics or ceramic coatings do not induce any cytotoxic effect.